Analysis of the molecular basis for mRNA dynamics in the nucleus using next generation sequencer and the function for the factor(s)of quality control.

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The mRNA quality control in the nucleus remains largely unknown. Here, to analyze the molecular function of mRNA quality control factors in the nucleus, whole transcriptome analysis of the mRNA dynamics in the nucleus was exploited. And the importance of the mRNA quality control was also investigated using the conventional biochemical and molecular biology methods. The whole transcriptome was analyzed from the cells treated with siRNA of RRP45, a core component of exosome, or MTR4, a subcomponent of exosome. Whole reads were mapped on the human genome and the changes of the number of reads mapped on the specific region were compared between siRNA treated- and control cells. The genes whose expressions were markedly increased wereselected by the treatment of siRNA of RRP45 or MTR4. Next, the identification and localization of human TRAMP (PAPD5-ZCCHC7-MTR4) complex were analyzed using conventional methods like immunoprecipitation and immunostaining. Exosome and MTR4 were localized in the nucleolus. PAPD5 and ZCCHC7 were localized in the nucleus. The candidate molecules of human TRAMP complex were associated in the nuclear extract.